Radio frequency (RF) only multipole spectrometers, more particularly quadrupole spectrometers, are widely applied in mass spectrometry and nuclear physics, due to their ability to transport ions with minimal losses. During such transportation of the ions, the initial ion positions and velocities change, but the total phase space volume occupied by the ion beam remains constant (see Dawson, P. H., “Quadrupole Mass Spectrometry and its Applications,” Elsevier Scientific Publishing Co., New York, 1976). However, if a buffer gas is introduced into the ion guide, a dissipative process occurs, due to ion molecule collisions, and this enables an ion beam to be focused onto the quadrupole axis after the initial velocities have been damped.
Collisional quadrupole or other multipole devices have been used as an ion guide providing an interface between an ion source and a mass spectrometer, or alternatively as a collision cell for collision-induced dissociation (CID) experiments. As a straightforward interface, collisional damping reduces the space and velocity distributions of the ions leaving the ion source, thus improving the beam quality. For CID experiments, primary ions having relatively large velocities enter the multipole and collide with buffer gas molecules, and so collision-induced dissociation takes place. The multipole helps to keep both primary ions and fragment ions, resulting from the collision-induced dissociation, close to the axis and to deliver them to the exit for further analysis. Collisions inside the multipole spectrometer again act to reduce the space and velocity distribution of the ion beam.
Ion motion in a perfect quadrupole field is governed by Mathieu's equation (See Dawson as cited above); ions oscillate around the quadrupole axis at an appropriate fundamental frequency which is determined by their m/z and quadrupole parameters, and is independent of ion position and velocity. If the frequency of any periodic forces acting on ions coincides with the ion fundamental frequency, then resonance excitation takes place. Similar resonance excitation is widely applied in quadrupole ion trap or in ion cyclotron resonance mass spectrometers (R. E. March, R. J. Hughes, “Quadrupole storage mass spectrometry,” 1989, John Wiley & Sons).
These properties of spectrometers have been employed in many ways. Thus, in U.S. provisional patent application 60/046,926 filed May 16, 1997 (and related U.S. patent application Ser. No. 09/066,556 and Canadian patent application 2,236,199), there is disclosed a high pressure MS-MS system. This was intended to provide improvements to a conventional triple quadrupole mass spectrometer arrangement, employing two precision quadrupole mass spectrometers separated by an RF-only quadrupole which is operated as a gas collision cell. The first mass spectrometer is used to select a specific ion mass-to-charge ratio (m/z), and to transmit the selected ions into the RF-only quadrupole or collision cell. In the RF-only quadrupole collision cell, some or all of the parent ions are fragmented by collisions with the background gas, commonly argon or nitrogen, at a pressure of up to several millitorr. The fragment ions, along with any unfragmented parent ions are then transmitted into the second precision-quadrupole which is operated in a mass resolving mode. Usually, the mass resolving mode of this second spectrometer is set to scan over a specified mass range, or else to transmit selected ion fragments by peak hopping, i.e. by being rapidly adjusted to select specific ion m/z ratios in sequence. The ions transmitted through this spectrometer are detected by an ion detector. A problem with this conventional arrangement is that the two mass resolving quadrupoles are required to operate in the high vacuum region (less than 10−5 torr), while the intermediate collision cell operates at a pressure up to several millitorr. That earlier invention was intended to simplify the apparatus and eliminate the necessity for separate RF-only and resolving spectrometers at the input to the apparatus. Instead, a single quadrupole is provided, operating in the RF-mode to act as a high pass filter. Additionally, this quadrupole is provided with an AC field, which can be identified as a “filtered noise field”, which contains a notch in the frequency range corresponding to the mass of an ion of interest. This notch can be moved, to select and separate desired ions.
Other older proposals can be found, for example, in U.S. Pat. No. 5,420,425 (Bier et al. and assigned to Finnigan Corporation). This relates to an ion trap mass spectrometer, for analyzing ions. It has electrodes shaped to promote an enlarged ion occupied volume. A quadrupole field is provided to trap ions within a predetermined range of mass to charge ratios. Then, the quadrupole field is changed so that trapped ions with specific masses become unstable and leave the trapping chamber in a direction orthogonal to the central axis of the chamber. The ions leaving the spectrometer are detected, to provide a signal indicative of their mass-to-charge ratios. One method that is taught in this patent is to first introduce ions within a predetermined range of mass-to-charge ratios into the chamber and subsequently to change the field to select just some ions for further manipulation. The quadrupole field is then adjusted so as to be capable of trapping product ions of the remaining ions, and the remaining ions are then dissociated or reacted with a neutral gas to form those product ions. Subsequently, the quadrupole field is changed again, to remove, for detection, ions whose mass-to-charge ratios lie within the desired range, which ions are then detected.
The above process describes how the technique of MS/MS (or MS2) is applied in an ion trap configuration. A related technique of MS/MS/MS (or MS3) can be provided by isolating one of the product ions of the first MS2 process, and eliminating all but the selected product ion from the trap. The selected product ion mass is then excited so that it fragments through collisions with the buffer gas in the trap. The range of secondary product ions formed in this two-stage process is then scanned from the trap for detection, so that a mass spectrum is recorded. The spectrum consists of fragments of a fragment from the original parent ion. The process can be extended by trapping and isolating one of the secondary product ions, and then fragmenting that ion mass, in order to form an MS/MS/MS/MS or MS4 spectrum, and ultimately the process can be extended to an MSn spectrum. Ion losses occur at each stage, however, so that sensitivity decreases as the number of steps increases. Nevertheless, this technique of MSn can be a useful tool to help elucidate the structure of organic ions.
Another approach to obtaining an MS3 spectrum is described in U.S. Pat. No. 6,011,259 by Craig Whitehouse, Thomas Dresch and Bruce Andrien of Analytica of Brantford, which shows how a multipole ion guide can be combined with a time-of-flight (TOF) mass spectrometer to provide MSn analysis. They describe the method of using resonant excitation to excite one ion mass in the quadrupole ion guide in order to fragment a selected ion (without isolating or rejecting the other ion masses). By turning the excitation on and off several times per second, a background subtracted spectrum of the fragments of the desired precursor ion can be created. This works very well with TOF mass spectrometers where the TOF is pulsed at a higher frequency than the pulsing of the excitation.
The method described produces an MS2 spectrum of the selected ion. In order to produce an MS3 spectrum (designated by the inventors of that patent as MS/MS2), two frequencies must be added, first exciting just the first precursor ion, then adding another frequency to excite both the first precursor ion and the selected product of that precursor ion together, and subtracting the spectra to obtain an MS3 spectrum. A total of three spectra must be collected sequentially: a first spectrum without any excitation, then a spectrum with only one excitation frequency (first MS2 spectrum), then a spectrum with both frequencies added simultaneously (MS3). The second spectrum must be subtracted from the first spectrum in order to generate the MS2 spectrum and identify the primary product ion of interest, and then the third spectrum must be subtracted from the second spectrum in order to generate an MS3 spectrum (in other words, an MS2 spectrum of the primary product ion).
The first technique taught above is complex, and requires a number of separate quadrupoles or the like, and the ability to move the ions sequentially through the different quadrupole sections. The technique taught in the Finnigan patent is complex and requires a number of steps. Also, it is concerned with ion traps and not a flow quadrupole. While all of the above methods can be used to obtain MS3 spectra (or higher order), they all suffer from some limitations or drawbacks. The ion trapping methods require isolation of the first parent ions before fragmentation, and then sequential steps of fragmentation, isolation and fragmentation in order to reach MS3. The initial ion mass which is fragmented is not mass selected.
The method described in Whitehouse et al to achieve MS3 is complex, and reduces the duty cycle for the overall process (as described by the inventors of the '259 patent) to 33% for MS3. Also, the mass-selective specificity of the first fragmentation step, obtained by exciting the ion radially to collide and fragment is much less than that achievable by using a mass spectrometer (a quadrupole mass filter for example). Therefore, the method taught is both less sensitive and less mass-selective than that described in the present application. Finally, these inventors fail to recognize that simple subtraction alone will not always give an accurate representation of the true fragment spectra; the present inventors have realized further statistical analysis is required to eliminate uncertainties due to poorly subtracted spectra.
Accordingly, it is desirable to provide one technique which, in one device, readily enables ions of a selected mass-to-charge ratio to be subject to collision-induced-dissociation (CID) or fragmentation, so that the fragments can be transported further for subsequent analysis. It is desirable to provide this in a single device, since movement of ions from one device to another inevitably leads to some losses. Similarly, the techniques of the Finnigan patent works effectively with pulse ion sources, but inefficiently with continuous ion flow, for instance from an electrospray ion source. In this field, spectrometers are frequently used to analyze small samples, and often, high efficiency is required, if any reliable reading or measurement is to be obtained.